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. 2025 Dec 11;17:751. doi: 10.1038/s41467-025-67438-5

Fig. 4. WAPL depletion in CTCF-Nm cells rescues distal deletional, but not inversional Vκ joins.

Fig. 4

a Top panel, diagram of the AID2 system for inducible degradation of WAPL. Bottom panel, schematic showing strategy of WAPL depletion in G1-arrested cells. Cells were first treated with 5-Ph-IAA for 4 h, followed by 4 days of STI-571 treatment to induce G1 arrest with continuous presence of 5-Ph-IAA. b Flow cytometry analysis demonstrating rapid degradation of WAPL after 4 h and persistent WAPL depletion in G1 cells upon 5-Ph-IAA treatment. The control parental line is the Rosa26-OsTIR1(F74G) v-Abl pro-B cell line. c Left, representative 3C-HTGTS analysis of untreated and 5-Ph-IAA treated Wapl-AID2 CTCF-Nm cells from Cer bait. I denotes the 3.2-Mb region upstream of the Igκ locus, II denotes the 3.2-Mb Igκ locus and III denotes the 3.2-Mb region downstream of the Igκ locus. Right, relative interactions of indicated regions are shown as percentage of the total read counts within the region encompassing from 3.2-Mb upstream of Igκ to 3.2-Mb downstream of Igκ. Data represents mean ± SD from three independent replicates. d Left, zoomed-in views show Igκ interaction profiles in untreated and 5-Ph-IAA treated Wapl-AID2 CTCF-Nm cells. Right, quantification of interactions showing median interaction counts of indicated regions using 50-kb bins. Data represents mean ± SD from three independent replicates. P values were calculated using unpaired, two-tailed Student’s t test. e Left, HTGTS-V(D)J-Seq analysis of Vκ usage frequency (%) in untreated (top) and 5-Ph-IAA treated Wapl-AID2 CTCF-Nm (bottom) v-Abl pro-B cells. Right, summary of deletional and inversional Vκ usage in proximal versus middle/distal Vκ regions in untreated and 5-Ph-IAA treated Wapl-AID2 CTCF-Nm cells. Data are shown as mean ± SD from three independent replicates. P values were calculated using unpaired, two-tailed Student’s t test.